1. Field of the Invention
The invention relates to crystalline TGF-.beta.2, which can be used as a starting material in any of the methods of use for TGF-.beta.2 outlined in the patents set forth below, but is particularly useful for the determination of the three-dimensional structure of the TGF-.beta.2 polypeptide.
The three dimensional structure of TGF-.beta.2 provides information which has a number of uses; principally related to the development of pharmaceutical compositions which mimic the action of TGF-.beta.2 or other members of the TGF-.beta. family of proteins.
2. Description of the Related Art
Transforming growth factor beta (TGF-.beta.) belongs to a family of growth factors that structurally related, which consists of at least five isoforms, TGF-.beta.1 through TGF-.beta.5, and activins and inhibins. These growth factors are involved in regulation of growth and function of many different cell types. TGF-.beta.s have been shown to stimulate proliferation of certain cells, such as osteoblasts and normal rat kidney fibroblasts grown on soft agar. However, TGF-.beta.s inhibit the proliferation of most cell types, such as lymphocytes, fibroblasts, endothelial cells and epithelial cells (1-4). In addition to its regulatory roles in cell proliferation, TGF-.beta.s are also known to regulate a variety of cellular processes, such as the production of extracellular matrix, chemotaxis, differentiation and morphogenesis. Due to the wide regulatory role of proteins of the TGF-.beta. family, there are a number of ongoing clinical studies involving the use of TGF-.beta.s, for example, to suppress inflammation, promote healing of soft tissue wounds, repair damaged bone and cartilage and to control tumor growth (5).
Despite the general multifunctional regulatory role of TGF-.beta.s, the mechanism by which TGF-.beta.s regulate cellular processes is unknown. Recently, significant advances have been made in understanding the roles of the three known types of TGF-.beta. receptors in signal transduction. A possibility has been raised that TGF-.beta. type II receptor may function as a serine/threonine kinase, based on homology of its amino acid sequence to other such kinase proteins, as a mechanism of signal transduction (6).
TGF-.beta.s are first expressed in cells as a precursor, which is then thought to be activated through proteolytic cleavage. The mature TGF-.beta. has 112 amino acids and forms a functional active homodimer of about 25 kilodaltons molecular weight. The amino acid sequences of TGF-.beta. are highly conserved among the human, bovine, porcine, simian, chicken and murine proteins. The high degree of structural conservation among the different species is consistent with the importance of TGF-.beta. in regulation of cellular functions.
One route to understanding the function of proteins is to take a structural point of view. Specifically, one determines the 3-D structure of a protein and inquires as to how the structure relates to function. In the case of TGF-.beta., the protein is actually one of a large family of similar proteins. It is also known that different forms of TGF-.beta. display slightly different affinities to their receptors and this differential binding onto the receptors is regulated according to tissue type, cell type which in turn regulates the cell proliferation. Thus it is critical to understand the structural differences between different forms of TGF-.beta.s in order to reveal their differences in biological function.
The following U.S. Patents, disclosing material relating to either compositions or methods of use for TGF-.beta. or TGF-.beta.2, have issued. All are hereby incorporated by reference.
______________________________________ U.S. Pat. No. issued subject matter ______________________________________ 4,434,094 2/28/84 partially purified TGF-.beta. 4,627,982 12/9/86 OFE, partially pure OIF 4,760,131 7/26/88 Collagen/heparin gel containing platelet releasates for wound healing 4,774,228 9/27/88 TGF-.beta.2 for promoting connective tissue deposition 4,774,322 9/27/88 pure TGF-.beta. 4,806,523 2/21/89 TGF-.beta. for immune modulation 4,816,442 3/2/89 TGF-.beta.2 inhibition of tumor growth 4,810,691 3/7/89 TGF-.beta.2 for promoting cell proliferation 4,843,063 6/27/89 TGF-.beta. for local/systemic bone repair 4,950,483 8/21/90 Collagen/heparain sponge con- taining a synergistic combination of TGF-.beta. and FGF 4,971,952 11/20/90 Method for treating inflammation 5,008,240 4/16/91 Method for treating inflammation 5,024,84 11/16/91 Collagen wound healing matrices and process for their production ______________________________________